Registration Dossier

Administrative data

Description of key information

The evaluation of the repeated dose toxicity was based on three studies:
- Two subchronic oral toxicity tests (rat) with mixtures containing a high concentration of DOT(2 -EHMA) (70 and 97% purity)- no guideline studies;
- One subchronic toxicity test (dog) with Dioctylin bis (IOMA) and dioctyltin bis (2-EHMA) which are isomers of the same compound and are structural analogues of each other. Based on the recently conducted developmental toxicity studies in two species it is considered that DOTI is likely to be more toxicoligically active and therefore use of data on this substance would be considered to be a worst case assessment of the registered substance.
No data on dermal or inhalatory repeated dose toxicity are available.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conduct prior to guideline and GLP implementation, however study is well documented and complies to a large extent to the later implemented guideline OECD 408 and GLP requirements. No information on the homogeneity or stability of the test substance in the diet was provided.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity in Rodents)
Deviations:
yes
Remarks:
some limitations
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source:no data
- Age at study initiation:no data
- Weight at study initiation:no data
- Fasting period before study:no data
- Housing: Test animals were housed in groups of 5
- Diet (e.g. ad libitum): not precised, ad libitum
- Water (e.g. ad libitum): not precised, ad libitum
- Acclimation period:no data

ENVIRONMENTAL CONDITIONS
- Temperature (°C):24
- Humidity (%):no data
- Air changes (per hr):no data
- Photoperiod (hrs dark / hrs light):no data
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
In the first experiment, groups of 30 weanling Wistar rats (CIVO-colony) (15 rats/sex/group) were exposed to the test substance, administered in the diet, at dose levels of 0, 100, 500, and 1000 ppm. Due to excessive mortality at 1000 ppm, a second experiment was initiated.
Groups of rats (15 per sex) were exposed to dose levels of 0, 50, and 250 ppm test substance in the diet. In a third experiment, groups of rats were exposed to 10 and 25 ppm of the test substance in the diet.

The test diets were prepared every two weeks and stored at room temperature. The test substance was thoroughly mixed in the stock diet.
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
90 days
Frequency of treatment:
continuously
Remarks:
Doses / Concentrations:
100, 500, and 1000 ppm (experiment 1)
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
50 and 250 ppm (experiment 2)
Basis:
nominal in diet
Remarks:
Doses / Concentrations:
10 and 25 ppm (experiment 3)
Basis:
nominal in diet
No. of animals per sex per dose:
15 rats/sex/group (experiment 1,2 and 3)
Control animals:
yes, concurrent no treatment
Details on study design:
no
Positive control:
no
Observations and examinations performed and frequency:
Body weights were recorded weekly. Food intake (by group) was determined during the exposure period. Food efficiency was calculated over the first four weeks of exposure. Water consumption (by group) was determined during the first week of exposure. Blood samples were collected in weeks 6 and 12.

Hematological data included hemoglobin content, packed cell volume, red blood cell counts, and total and differential white blood cell counts. Biochemical data collected terminally included blood sugar, blood urea nitrogen (BUN), serum glutamic-pyruvic transaminase (SGPT), serum glutamic oxaloacetic transaminase (SGOT), serum alkaline phosphatase (SAP), and total serum protein (TSP).
Kidney function was examined by determining glutamic oxaloacetic transaminase (UGOT) in the urine and urine protein.
Urine samples were collected, pooled by treatment level, and examined for appearance, pH, glucose, albumin, occult blood, ketones, and microscopy of the sediment.

Sacrifice and pathology:
At the end of the experimental period, animals were sacrificed and examined macroscopically for pathological changes. In the first experiment, the water content of the brain was determined in 10 rats (5 per sex) from each treatment group. In the second experiment, the tin content of pooled samples of the liver, kidneys, and testicles of 10 rats of each treatment group was determined.

Detailed microscopic examination of organs and tissues in the first experiment (control and 500 ppm groups) included: lungs, trachea, pancreas, salivary glands (parotid, submaxillary, and sublingual), prostate, epididymis, uterus, coagulating gland, seminal vesicle, thoracic aorta, stomach, duodenum, ileum, cecum, colon, mesenteric and axillary lymph nodes, urinary bladder, skeletal muscle, spinal cord, femoral nerve, sternum with marrow, exorbital lachrymal glad, preputial gland, and skin. Microscopic examination of rats in the 100 ppm group was limited to the liver, kidneys, spleen, thymus, lymph nodes, heart, and testicles. Animals that died before the end of the exposure period were not examined pathologically.
In the second experiment, microscopic examinations of all groups were limited to the liver, kidney, spleen, thymus, and lymph nodes.
Other examinations:
no
Statistics:
no data
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
effects observed, treatment-related
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
no effects observed
Details on results:
Mortality occurred at 250, 500, and 1000 ppm.

Mean body weights and food intake were reduced at 250, 500, and 1000 ppm.

Hemoglobin content and white blood cell counts were slightly decreased at 250 and 500 ppm. There was a considerable increase in the number of neutrophils and leukocytes, and a decrease in the number of lymphocytes at >=100 ppm.

There were minimal to moderate changes in the microscopic findings in urine. Specific gravity of the urine was decreased and UGOT levels were increased at 500 ppm. Specific gravity of the urine of females at 100 ppm was decreased. SAP levels were significantly increased at >=100 ppm. Glucose levels were decreased at 500 ppm and BUN levels were increased in males at 500 ppm. SGPT levels for males and females were increased at 500 and 100 ppm, respectively.

There was no increase in water content of the brain. Relative liver and kidney weights were slightly increased at >=100 ppm. Relative thymus weights were reduced at >=25 ppm. The relative weights of the heart (females), brain (both sexes), and gonads (males) were significantly increased at 500 ppm. The increase in relative weight of the liver was not dose-dependent. The relative weights of the brain and the testicle showed an inverse relationship with body weights. The relative spleen weight was slightly increased in some dose groups. Treatment-related gross pathological changes observed included a reduction in thymus size at >=100 ppm. The number of lymphocytes in the
thymus were depleted at >=100 ppm.

Other changes observed included signs of chronic respiratory disease, unilateral hydronephrosis, and proteinaceous plugs in the urinary bladder; however, the latter are commonly found in the strain of rats tested and, therefore, may not be related to exposure to the test substance. Treatment-related histological lesions in the liver and kidneys ranged from minimal (250 ppm) to moderate (500 and 1000 ppm).

The no-observable-adverse-effect-level (NOAEL) was determined to be 10 ppm in the diet (equivalent to 0.5 mg/kg/bw/day) of rats exposed for 90 days, on the basis of reduced thymus weight at 25 ppm. Based on these findings, the lowest-observable-adverse-effect level (LOAEL) was determined to be 25 ppm in the diet (calculated as 1.07-1.24 mg/kg bw/day in males and 1.46-1.51 mg/kg bw/day in females). Calculation of dosage was performed using body weights of 0.34 kg (males) and 0.2 kg (females), and average food consumption of 14.6-16.8 g/rat/day (males) and 11.7-12.1 g/rat/day (females).
Dose descriptor:
NOAEL
Effect level:
10 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: reduced thymus weight (10 ppm is equivalent to 0.5 mg/kg/bw/day)
Critical effects observed:
not specified
Conclusions:
The NOAEL for DOT(2-EHMA) was determined to be 10 ppm (equivalent to 0.5 mg/kg bw/day) based on reduced thymus weight.
Executive summary:

A repeated dose oral toxicity study was carried out with rats and a mixture of Dioctyltin bis(2-EHMA) [CAS No. 15571-58-1]:Octyltin tris(2-EHMA) [CAS No. 27107-89-7]: Trioctyltin (2-EHMA) [CAS No. 61912-55-8] (97.0:0.3:2.17% mixture). The study predates GLP, but is well documented and quality assurance is comparable. No information on the homogeneity or stability of the test substance in the diet was provided, due to which the study was considered less reliable. The NOAEL was determined to be 10 ppm (equivalent to 0.5 mg/kg bw/day) and the LOAEL was determined to be 25 ppm (equivalent to 1.3 mg/kg bw/day) based on reduced thymus weight.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
0.5 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
The key study is a 90 day study in rats dosed via the diet; the study was conducted with a mixture containing the test material however it pre-dates GLP and standardised guidelines, however study is well documented and complies to a large extent to the later implemented guideline OECD 408 and GLP requirements. The key study was therefore assigned a reliability score of 2 in line with the criteria of Klimisch (1997). Two supporting studies are included; one is a 13 week study in rats which was conducted with a mixture containing the test material, the other is a 14 week study in dogs, conducted with the read-across substance Dioctylin bis (IOMA) and dioctyltin bis (2-EHMA). Both studies were conducted via the dietary route and both studies pre-date GLP and standardised guidelines. Both studies were assigned a reliability score of 2.
The overall quality of the database is good.

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

An evaluation of the repeated dose toxicity of the substance was based on the findings of three studies, as follows.

In the key study (anonymous, 1970) repeated dose oral toxicity of a mixture of Dioctyltin bis(2-EHMA) [CAS No. 15571-58-1]:Octyltin tris(2-EHMA) [CAS No. 27107-89-7]: Trioctyltin (2-EHMA) [CAS No. 61912-55-8] (97.0:0.3:2.17% mixture) was evaluated in rats dosed continuously via the diet at concentrations of 10, 25, 50, 250, 100, 500 and 1000 ppm. Fifteen rats per sex received each dose for a period of 90 days. Body weights, food and water consumption were recorded and blood samples were collected in weeks 6 and 12 for haematological and clinical chemistry evaluation. At the end of the experimental period, animals were sacrificed and subject to pathological and histopathological examination.

Under the conditions of the study the no-observable-adverse-effect-level (NOAEL) was determined to be 10 ppm in the diet (equivalent to 0.5 mg/kg/bw/day) of rats exposed for 90 days, on the basis of reduced thymus weight at 25 ppm.

In the supporting study (anonymous, 1974) repeated dose oral toxicity of Dioctyltin bis(2-EHMA) [CAS No. 15571-58-1]:Octyltin tris(2-EHMA) [CAS No. 27107-89-7] (70:30% mixture) was evaluated in rats dosed continuously via the diet at concentrations of 25, 50 and 100 ppm. Twenty rats per sex received each test diet for a period of 90 days. Blood and urine samples for haematology and biochemistry were collected during weeks 5, 9, and 13. Clinical symptoms of toxicity were recorded daily, and body weights and food consumption were recorded weekly. An ophthalmic examination was conducted pre-exposure and in rats from the 25 and 100 ppm dose groups, during weeks 5, 9, and 13.The following organs were weighed at necropsy: adrenals, kidneys, brain, liver, heart, gonads, and thymus.  

The following organs and tissues were collected for histopathological examination: adrenals, brain, gonads, kidneys, lymph nodes, muscle,  pituitary, spleen, thymus, aorta, colon, gross lesions, liver, pancreas, prostate, uterus, spinal cord, thyroids, bone marrow,  eye, heart, lungs, mammary gland, sciatic nerve, small intestine, stomach, and urinary bladder.

No deaths occurred and no clinical symptoms of toxicity were recorded. Body weight gains, food consumption, clinical chemistry, and urine analysis for treated rats were comparable to the control group. Ophthalmic examination did not reveal any abnormalities in treated rats. There was a significant dose-related reduction in absolute and relative thymus gland weights in the 50 ppm (3.3 mg/kg) and 100 ppm (6.6 mg/kg) dose groups.

The no effect level for DOT(2-EHMA) was determined to be 25 ppm (calculated as 1.25 mg/kg/day, based on a food factor of 0.05) based on reduced absolute and relative thymus gland weights.

In the supporting study reported by Johnson (1970) repeated dose toxicity the structural analogue Di(n-octyl) tin S, S'-bis (isooctylmercaptoacetate), was evaluated in dogs dosed continuously via the diet at concentrations of 0, 20, 50, and 150 ppm. Three dogs per sex received each test diet for a period of 14 weeks. The animals were observed for mortality, body weight, general physical condition and weekly physical examinations, blood pressure, electrocardiograms and eye examinations. Haematology and clinical chemistry determinations were performed at weeks 0, 4, and 13. The dogs were sacrificed after 14 weeks and subjected to gross pathological and histopathological assessment.

All animals survived the duration of the experiment and maintained good physical condition throughout. There were no treatment related effects on body weights, food consumption, urinalysis, haematology, clinical chemistry and ophthalmic examinations were negative except for one animal. No gross pathological or histopathological changes were noted.

Therefore, under the conditions of the study the NOAEL was determined to be 150 ppm, the highest dose level tested.

 

Findings from the study reported by Appel (2004) are included in the dataset since the test material was previously regarded as being an adequate substance for read-across to the registered substance since DOT(2-EHMA) had previously been demonstrated to be readily hydrolysed to Dichlorodioctyltilstanane (CAS no.3542-36-7) under physiological conditions (see section 7.1.1). Thus DOTC(Dichlorodioctylstannane) was considered to be an appropriate anchor compound and surrogate for the mammalian toxicology endpoints of repeated dose, in vivo genetic toxicity, reproduction and developmental effects, when it is dosed via the oral route of administration.

However, read-across to the substance DOTC is no longer considered as wholly appropriate based on the results of the recent Hydrolysis studies, as reported by Naßhan H 2014 and Naßhan, H, 2015 (see section 7.1.1) which indicate the substance DOTECl is the only metabolite of DOTE which is formed in a simulated mammalian gastric environment; no dioctyltindichloride was formed under the conditions of the study. Findings from the study reported by Appel (2004) have therefore been disregarded and are not included in the overall assessment of repeated dose toxicity of the registered substance.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
The key study was selected as such as it is the study in which the test material most closely relates to the registered substance. Findings from this study are therefore deemed to be of most relevance and reliability for assessment of the registered substance.
The key study also reports the lowest NOAEL and can therefore be regarded as the most precautionary result to use for risk assessment.

Repeated dose toxicity: via oral route - systemic effects (target organ) cardiovascular / hematological: thymus

Justification for classification or non-classification

Based on the available data on the substance it self ( pure and mixture) and on the gastric hydrolysis product (pure DOTC 94%), DOT(2-EHMA) is classified as T: R48/25 according to Directive 67/548/EEC and according to CLP : STOT RE category 1 for specific target organ toxicity, specifically for effects on the thymus (H372).

Justification: A LOAEL is 0.7 mg/kg bw/d. This value is smaller than 10, therefore a classification 'STOT RE 1' is required according to the regulation EC no.1272/2008 (CLP).

This value is smaller than 5 mg/kg bw/d, therefore according to the Directive 67/548/EEC, the substance is classified "R48 /25" (with a LOAEL between 5 and 50 mg/kg bw/d : R48 /22).